Compositions and methods for treating alopecia

ABSTRACT

The present invention is directed to a composition for treating alopecia containing minoxidil, cyclosporine A and a compound that binds FK506 binding protein 4 suitable for administration to humans. The present invention is further directed to treating alopecia in humans by administering a composition of the invention.

FIELD OF THE INVENTION

The present invention is directed to a composition for treating alopeciacontaining a compound that binds FK506 binding protein 4 suitable foradministration to humans. The present invention is further directed totreating alopecia in humans by administering a composition of theinvention.

BACKGROUND OF THE INVENTION

Androgenic alopecia (i.e. male pattern baldness) poses seriouspsycho-social issues for millions of effected individuals. Theseindividuals include 54% of all US men aged 30 or more and 50 to 75% ofUS women over the age of 65. Otberg, N., et al., Androgenetic alopecia,Endocrinology and metabolism clinics of North America, 2007 June, 36(2),379-398 and Scheinfeld, N., A review of hormonal therapy for femalepattern (androgenic) alopecia, Dermatology online journal, 2008. 14(3),1-5. This translates to more than 150 million individuals in the USalone.

The pathophysiology of androgenic alopecia is not well understood, butas the name implies, testosterone plays a key role in male patternbaldness and hair loss. This role was inferred from the finding thatcastrated men do not suffer from hair loss. Otberg et al., 2007.Similarly, pseudohermaphrodites that are null at the 5-alpha reductase(“5-AR”) loci, which encode for an enzyme involved in testosteronemetabolism, are also protected from androgenic alopecia. Ellis J. A., etal., Male pattern baldness: current treatments, future prospects, DrugDiscovery Today, 2008, 13, 791-797. Testosterone, a lipophilic moleculethat diffuses the cell membrane, is converted into its more active form,dihydrotestosterone (“DHT”), by cytoplasmic 5-AR. There are two types of5-AR, 1 and 2, with type 2 5-AR found in the skin and the inner rootsheath of hair follicles. Burkhart C. G., et al., 5 alpha-reductase andfinasteride in pattern alopecia and acne, Journal of Drugs inDermatology, 2004, 3, 363-364. Once DHT enters the nucleus it binds tothe androgen receptor, regulating gene expression. However, the genesinvolved in mediating male pattern baldness have yet to be identified.

Chemotherapy can also lead to alopecia including permanentandrogenic-like alopecia. Kluger N et al., Permanent scalp alopeciarelated to breast cancer chemotherapy by sequentialfluorouracil/epirubicin/cyclophosphamide (FEC) and docetaxel: aprospective study of 20 patients, Ann Oncol. 2012 November, 23(11),2879-84, Epub 2012 May 9. Some chemotherapeutic agents that have beenassociated with permanent alopecia include docetaxel, anthracycline andcyclophosphamide, carboplatin and trastuzumab, paclitaxel and busulfan.Basilio F M et al., Clinical and histological study of permanentalopecia after bone marrow transplantation, An Bras Dermatol, 2015November-December, 90(6), 814-21. To date the mechanism forchemotherapy-induced permanent alopecia is unknown.

Only minoxidil, a topically applied compound, and finasteride, an oralmedication, have been approved to treat androgenic alopecia. Minoxidilwas originally developed as a systemic vasodilating agent to treathypertension, however many patients suffered with disseminatedhypertrichosis as a result of treatment. Bienova M., et al, Androgeneticalopecia and current methods of treatment, Acta DermatovenerologicaAlpina, Pannonica, et Adriatica, 2005, 14, 5-8. It was soon discoveredthat topical application of minoxidil results in limited hairrestoration, largely confined to the sites of application. Analysis ofthe cellular and molecular mechanisms of minoxidil-mediated hair growthhas shown that it promotes the survival of dermal papilla cells (“DPCs”)of human hair follicles, by activating both ERK and Akt and bypreventing cell death by increasing the ratio of Bcl-2/Bax. Han J. H.,et al., Effect of minoxidil on proliferation and apoptosis in dermalpapilla cells of human hair follicle, J Dermatol Sci, 2004, 34, 91-98.The DPCs are thought to provide trophic support to the hair follicle.Rendl M., et al., BMP signaling in dermal papilla cells is required fortheir hair follicle-inductive properties, Genes Dev 2008, 22, 543-557.Among the proteins up-regulated by minoxidil are the trophic factorsvascular endothelial growth factor (“VEGF”), hepatocyte growth factor(“HGF”), insulin-like growth factor 1(“IGF-1”) and bone morphogenicprotein 4 (“BMP-4”). Ryu S. et al., Mycophenolate antagonizesIFN-gamma-induced catagen-like changes via beta-catenin activation inhuman dermal papilla cells and hair follicles, International Journal ofMolecular Sciences 2014, 15, 16800-16815. Further, minoxidil is known topotentiate HGF and IGF-1 actions through the activation of uncoupledsulfonylurea receptor (“SUR”) on the plasma membrane of DPCs. Minoxidilhas been shown to be effective in maintaining existing hair follicles,but ineffective in stimulating new follicles. Sinclair has shown thatonly 15% of those treated with minoxidil had new hair growth, while 50%of those treated maintained existing hair, with no additional loss at6-months. Sinclair R., Male pattern androgenetic alopecia, BMJ, 1998,317, 865-869. Notably, discontinuation of minoxidil treatment results inthe resumption of hair loss, presumably through the loss of trophicsupport.

Finasteride, unlike minoxidil, is an oral medication, with potentiallysevere side effects including erectile dysfunction, gynecomastia, andloss of libido. Finasteride is a competitive 5-AR inhibitor thatinhibits the conversion of testosterone to DHT, resulting in a decreasein androgenic alopecia. Price V. H., Treatment of hair loss, The NewEngland Journal of Medicine, 1999, 341, 964-973.

In addition to minoxidil and finasteride, cyclosporine A (“CSA”), animmunosuppressive drug intended to prevent rejection of solid organallografts, promotes robust hair growth in up to 80% of transplantpatients receiving systemic treatment. Wysocki G. P., et al.,Hypertrichosis in patients receiving cyclosporine therapy, Clin ExpDermatol, 1987, 12, 191-196. While CSA is highly effective in preventinggraft rejection, it has severe and undesirable side effects when takenorally or parenterally. Thus, CSA is a poor choice for systemicadministration in all but the most life threatening situations. Incontrast, the topical administration of low-dose, topical CSA is notassociated with immune suppression, hypertension, renal toxicity or theother severe or life-threatening side effects seen with oral CSAadministration.

The mechanism of CSA-mediated hair growth was recently elucidated byFuchs and her colleagues. These workers demonstrated that NFATc1 isexpressed preferentially by the follicular stem cell, where it acts torepress stem cell proliferation. Upon administration, CSA binds to thecalmodulin-dependent, serine/threonine protein phosphatase calcineurin,which in turn binds to NFAT, and relieves the repression on thefollicular stem cell. With the NFAT repression relieved, the follicularstem cells proliferate, resulting in precocious follicular growth.Horsley V., et al., NFATc1 balances quiescence and proliferation of skinstem cells, Cell, 2008, 132, 299-310. Interestingly, as opposed to oraladministration, topical treatment with CSA has shown to be veryinefficient at promoting hair growth in androgenic alopecia when usedalone, or in combination with minoxidil. Gilhar A., et al., Topicalcyclosporine in male pattern alopecia, J Am Acad Dermatol, 1990, 22,251-253; Buhl A. E., et al., Differences in activity of minoxidil andcyclosporin A on hair growth in nude and normal mice. Comparisons of invivo and in vitro studies, Laboratory Investigation; A Journal ofTechnical Methods and Pathology, 1990, 62, 104-107.

Another compound that has shown hair growth stimulation is RT175(AMG-474-00, GM1485, GPI 1485). RT175 is a 241 Dalton molecule havingthe following chemical structure

RT175 has been shown to re-grow hair in rats that have undergonecraniotomy prior to neurosurgery. Ducruet et al., GM1485, anonimmunosuppressive immunophilin ligand, promotes neurofunctionalimprovement and neural regeneration following stroke, J Neurosci Res,2012 July, 90(7):1413-23. RT175 has also been shown to induce hairgrowth in shaved mice. European Patent No. 1842845 to GliaMed, Inc.,published Oct. 10, 2007. RT175 binds with high affinity to FK506 bindingprotein 4 (“FKBP52”). FKBP52 is known to act as a molecular chaperonefor the glucocorticoid receptor (“GR”). After binding to ligand, theRT175/GR complex translocates to the nucleus. Banerjee A., et al.Control of glucocorticoid and progesterone receptor subcellularlocalization by the ligand-binding domain is mediated by distinctinteractions with tetratricopeptide repeat proteins, Biochemistry, 2008,47, 10471-10480. It has been shown that that RT175 treatment offibroblasts for 2 hours results in the translocation of FKBP52 to thenucleus, presumably with its cargo. This timeframe is consistent withcDNA array studies that have shown a significant upregulation ofchromatin remodeling via the transcription factors Ikaros and Afql,suggesting that treatment with RT175 results in alteration in chromatinstructure as the cells are reprogrammed, including the upregulation ofsecreted molecules that are associated with hair growth, wnt1 and IGF2.Keshava C., et al., Transcriptional signatures of environmentallyrelevant exposures in normal human mammary epithelial cells:benzo[a]pyrene, Cancer Letters, 2005, 221, 201-211; Koipally J., et al.Ikaros chromatin remodeling complexes in the control of differentiationof the hemo-lymphoid system, Cold Spring Harb Symp Quant Biol 1999, 64,79-86; Galbiati F., et al., Caveolin-1 expression inhibitsWnt/beta-catenin/Lef-1 signaling by recruiting beta-catenin to caveolaemembrane domains, J Biol Chem, 2000, 275, 23368-23377; Tamimi Y. et al.,WNT5A is regulated by PAX2 and may be involved in blastemal predominantWilms tumorigenesis, Neoplasia, 2008, 10, 1470-1480; Nakao K., et al.,IGF2 modulates the microenvironment for osteoclastogenesis, BiochemBiophys Res Commun, 2009, 378, 462-466; Sun Y., et al., Evolutionarilyconserved transcriptional co-expression guiding embryonic stem celldifferentiation, PLoS ONE, 2008, 3, e3406; Andl T., et al., WNT signalsare required for the initiation of hair follicle development,Developmental Cell, 2002, 2, 643-653.

Despite the commercial availability of minoxidil and finasteride anddemonstrations of hair growth following systemic administration ofcyclosporine A and topical administration of RT175 in rodents thereremains a need in the art for a topical composition for the treatment ofalopecia including chemotherapy-induced alopecia in humans that avoidsthe side effects of oral or parenteral administration and islong-lasting.

SUMMARY OF THE INVENTION

In one aspect, the present invention is directed to a composition forthe treatment of alopecia comprising a compound that binds FK506 bindingprotein 4.

In another aspect, the present invention is directed to a compositionfor the treatment of alopecia comprising a compound that binds FK506binding protein 4 and one or more additional active agents selected fromthe group consisting of minoxidil, cyclosporine A, and a combinationthereof.

In another aspect, the present invention is directed to a compositionfor the treatment of alopecia comprising a compound of formula (I)

or a pharmaceutically acceptable salt or ester thereof, minoxidil andcyclosporine A, wherein R¹ is COOH, a methoxy, a phenyl, a benzyl, asubstituted phenyl or a substituted benzyl.

In a preferred embodiment the substituted phenyl and substituted benzylof the compound of formula (I) are each individually substituted with analkyl group, a methoxy group or a halogen.

In a more preferred embodiment the compound of formula (I) is selectedfrom the group consisting of

and a pharmaceutically acceptable salt or ester thereof.

In a most preferred embodiment the compound of formula (I) is RT175.

In another aspect, the present invention is directed to a compositionfor the treatment of alopecia comprising a compound of formula (II)

or a pharmaceutically acceptable salt or ester thereof, minoxidil andcyclosporine A.

In another more preferred aspect, the compositions of the presentinvention further comprise one or more excipients selected from thegroup consisting of urea, polyoxyl 40 stearate, a carbomer, cetylalcohol, glyceryl monostearate, mineral oil, ethanol, propylene glycol,polyglycol 300, citric acid, sodium phosphate dibasic, stearyl alcohol,isopropyl myristate, sodium hydroxide, petroleum jelly, xanthan gum,white petrolatum, sorbitol solution, cetearyl alcohol, ceteareth-20,simethicone, sodium benzoate, glyceryl monostearate, polyethylene glycolmonostearate, sorbic acid, butylated hydroxytoluene and water.

In a preferred embodiment the one or more excipients are a combinationof white petrolatum, sorbitol solution, propylene glycol, cetearylalcohol, ceteareth-20, simethicone, glyceryl monostearate, polyethyleneglycol monostearate, sorbic acid and butylated hydroxytoluene.

In another preferred embodiment, the one or more excipients are acombination of urea, polyoxyl 40 stearate, propylene glycol, polyglycol300 (Medibase C available from Medisca), citric acid, sodium phosphatedibasic, cetyl alcohol, stearyl alcohol, isopropyl myristate, sodiumbenzoate and water.

In a more preferred embodiment, the one or more excipients are acombination of about 1,200 grams of urea, about 103 grams polyoxyl 40stearate, about 63 milliliters of propylene glycol, about 47 millilitersof polyglycol 300, about 1 gram of citric acid, about 2 grams of sodiumphosphate, about 94 grams of cetyl alcohol, about 200 grams of stearylalcohol, about 219 grams of isopropyl myristate, about 3 grams of sodiumbenzoate and about 1,000 to about 1,500 milliliters of water.

In a preferred aspect, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration from about 1% to about 10% w/v, cyclosporine A at aconcentration from about 0.01% to about 1% w/v and RT175 at aconcentration from about 0.000001% to about 0.0001% w/v.

In another preferred aspect, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration from about 1% to about 10% w/v, cyclosporine A at aconcentration from about 0.01% to about 1% w/v, RT175 or apharmaceutically acceptable salt or ester thereof at a concentrationfrom about 0.000001% to about 0.0001% w/v, ethanol at a concentrationfrom about 10% to about 50% w/v, propylene glycol at a concentrationfrom about 10% to about 70% w/v and water at a concentration from about10% to about 50% w/v.

In a more preferred aspect, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v and RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v.

In another more preferred aspect, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v, RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v, ethanol at aconcentration of about 28% w/v, propylene glycol at a concentration ofabout 47% w/v and water at a concentration of about 19% w/v.

In another aspect, the present invention is directed to a method oftreating alopecia comprising topically administering to a human in needthereof an effective amount of a composition of the present invention.

In a preferred aspect, the present invention is directed to a method oftreating androgenic alopecia comprising topically administering to ahuman in need thereof an effective amount of a composition of thepresent invention.

In another aspect, the present invention is directed to a method oftreating alopecia in a human in need thereof comprising topicallyadministering concurrently or sequentially minoxidil, cyclosporine A anda compound that binds FK506 binding protein 4.

In another aspect, the present invention is directed to a method oftreating chemotherapy-induced alopecia comprising topicallyadministering to a human in need thereof an effective amount of acomposition of the present invention.

In a preferred aspect, the human in need of alopecia treatment suffersfrom achromotrichia and the method provides regrowth of pigmented hair,preferably the achromotrichia is due to aging.

In another aspect, the present invention is directed to a method ofenhancing facial hair (including eye brow) growth comprising topicallyadministering to a human in need thereof an effective amount of thecompositions of the present invention.

In a preferred aspect, the present invention is directed to a method oftreating alopecia in a human in need thereof comprising topicallyadministering concurrently or sequentially minoxidil at a concentrationfrom about 1% to about 10% w/v, cyclosporine A at a concentration fromabout 0.01% to about 1% w/v and RT175 or a pharmaceutically acceptablesalt or ester thereof at a concentration from about 0.000001% to about0.0001% w/v.

In a more preferred aspect, the present invention is directed to amethod of treating alopecia in a human in need thereof comprisingtopically administering concurrently or sequentially minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v and RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v.

In a more preferred aspect, the present invention is directed to amethod of treating alopecia in a human in need thereof comprisingtopically administering to a human in need thereof an effective amountof the composition comprising minoxidil, cyclosporine and RT175 or apharmaceutically acceptable salt or ester thereof, and urea.

In another aspect the present invention is directed to a method oftreating alopecia in a human in need thereof comprising the steps of:

-   -   (i) administering fractional laser treatment to an affected area        of a human; and    -   (ii) topically administering a composition of the invention,        wherein the steps can be in any order.

In another aspect the present invention is directed to a method oftreating alopecia in a human in need thereof comprising the steps of:

-   -   (i) pretreating an area affected with alopecia with 40% w/v        urea; and    -   (ii) topically administering a composition of the invention.

In another aspect, the present invention is directed to a method oftreating alopecia comprising topically administering concomitantly orsequentially a compound that binds FK506 binding protein 4 and at leastone compound selected from the group consisting of minoxidil andcyclosporine A.

In another aspect, the present invention is directed to a method oftreating alopecia comprising topically administering concomitantly orsequentially a compound that binds FK506 binding protein 4, minoxidiland cyclosporine A.

In another aspect, the present invention is directed to a method oftreating alopecia comprising topically administering to a female humanin need thereof an effective amount of a composition comprising acompound that binds FK506 binding protein 4, minoxidil; and cyclosporineA.

In another aspect, the present invention is directed to a method ofenhancing eyebrow growth comprising topically administering to a humanin need thereof an effective amount of a composition comprising acompound that binds FK506 binding protein 4, minoxidil; and cyclosporineA.

BRIEF DESCRIPTION OF THE DRAWINGS

The patent or application file contains at least one drawing executed incolor. Copies of this patent or patent application publication withcolor drawing(s) will be provided by the Office upon request and paymentof the necessary fee.

FIG. 1. Punch biopsies of mouse dorsal skin treated topically withRT175.

FIG. 2. Scalp closure following craniotomy in rats treated topicallywith RT175.

FIG. 3. Skin lesions in pigs treated topically with RT175.

FIG. 4. 59-year-old male with androgenic alopecia treated topically withRT175 and RT175/minoxidil.

FIG. 5. 57-year-old male with androgenic alopecia treated topically withRT175/minoxidil/cyclosporine A.

FIG. 6. 62-year-old female with androgenic alopecia treated topicallywith RT175/minoxidil/cyclosporine A.

FIG. 7. 57-year-old male with androgenic alopecia and achromotrichiatreated topically with RT175/minoxidil/cyclosporine A.

FIG. 8. 57-year-old male with chemotherapy-induced alopecia treatedtopically with RT175/minoxidil/cyclosporine A.

FIG. 9. 24-year-old male with inadequate facial hair growth treated withRT175/minoxidil/cyclosporine A.

FIG. 10. 45-year-old female with inadequate eyebrow growth treated withRT175/minoxidil/cyclosporine A.

FIG. 11. Skin regeneration and hair regrowth following shaving andrasping of mice dorsal skin and 10-day treatment with RT1061.

DETAILED DESCRIPTION OF THE INVENTION

Applicants unexpectedly discovered a composition for the treatment ofalopecia which has reduced side effects and prolonged effects overavailable treatments.

As used herein, “minoxidil” refers to the compound of the formula,

and any pharmaceutically acceptable salt or ester thereof.

As used herein, “cyclosporine A” refers to the compound of the formula,

and any pharmaceutically acceptable salt or ester thereof.

As used herein, “RT175” refers to the compound of the formula,

As used herein, “alopecia” refers to the loss of hair from the body,whether on the scalp, body, face or eyebrows, and due to a pathologicalcondition.

As used herein the term “pharmaceutically acceptable” refers toingredients that are not biologically or otherwise undesirable in atopical application.

As used herein the term “effective amount” refers to the amountnecessary to treat a patient in need thereof.

As used herein, “androgenic alopecia” refers to an autosomal disorderwhich begins in puberty in genetically disposed individuals. Androgenicalopecia is also known as hereditary baldness, male pattern baldness,and seborrheic alopecia. Androgenic alopecia may occur in males andfemales.

As used herein, “chemotherapy-induced alopecia” refers to hair loss thatbegins to occur from sometime after onset of chemotherapy to within onemonth of stopping of chemotherapy.

As used herein, “permanent chemotherapy-induced alopecia” refers tochemotherapy-induced alopecia wherein hair has not begun to regrowwithin 1 year of stopping chemotherapy.

As used herein, “achromotrichia” refers to an absence or loss of pigmentin the hair shaft. Achromotrichia may be due to aging, stress, diet ordisease.

As used herein, “fractional laser therapy” or “fractional lasertreatment” refers to application of a laser beam that is divided intothousands of zones and is capable of reacting with both the epidermisand dermis. This treatment is sometimes referred to as fractional laserphotothermolysis. Fractional lasers may be based on, but are not limitedto, erbium, carbon dioxide, diode, yttrium aluminum garnet (YAG),neodymium-doped yttrium aluminum garnet, yttrium scandium gallium garnet(YSGG) or combinations thereof. Commercial examples of fractional laserssufficient for the present invention include, but are not limited to,Profractional (Sciton, Inc.), Halo (Sciton, Inc.), Emerge (CynosurePalomar), Lux1540 (Cynosure Palomar), Lux2940 (Cynosure Palomar), DeepFX (Lumenis), Active FX (Lumenis), Fraxel re:pair® (Fraxel re:pair is aregistered trademark of Reliant Technologies LLC; available throughSolta Medical), Fraxel re:store® (Fraxel re:store is a registeredtrademark of Reliant Technologies LLC; available through Solta Medical),Clear+Brilliant (Solta Medical), Fraxel® Dual 1550/1027 (Fraxel® is aregistered trademark of Reliant Technologies LLC; available throughSolta Medical) and Pearl® Fractional (Pearl is a registered trademark ofCutera, Inc.).

As used herein, “treatment” or “treating” refers to preventing hair lossand growing, regrowing and regenerating hair. As used herein the term“pharmaceutically acceptable” refers to ingredients that are notbiologically or otherwise undesirable in a topical application.

As used herein the term “effective amount” refers to the amountnecessary to treat a patient in need thereof.

As used herein “% w/v” and “percent w/v” refer to the percent weight ofthe total formulation.

As used herein the term “R¹” refers to a substituent selected from thegroup consisting of COOH, a methoxy, a phenyl, a benzyl, a substitutedphenyl and a substituted benzyl.

In general, the term “substituted” means that one or more hydrogens ofthe designated moiety are replaced with a suitable substituent.

As used herein the term “alkyl” refers to a branched or straight-chainalkyl consisting of a saturated hydrocarbon group of 1 to 24 carbonatoms (C₁-C₂₄) unless otherwise stated. The alkyl group can be cyclic oracyclic. As used herein, all numerical values relating to amounts,weights, and the like, that are defined as “about” each particular valueis plus or minus 10%. For example, the phrase “about 10% w/v” is to beunderstood as “9% to 11% w/v.” Therefore, amounts within 10% of theclaimed value are encompassed by the scope of the claims.

In some embodiments, compositions of the present invention may contain asolvent. Solvents of the present invention include, but are not limitedto, ethanol, propylene glycol, water, polyethylene glycol, glycerol,isostearic acid, oleic acid, trolamine, tromethamine, triacetin,sorbitan monolaurate, sorbitan monooleate, sorbitan monopalmitate,butanol, iso-amyl acetate, methanol, propanol, isobutene, pentane,hexane, chlorobutanol, turpentine, cytopentasiloxane, cyclomethicone,methyl ethyl ketone and mixtures thereof. Total solvents of the presentinvention may be at concentrations from about 10% to about 99% w/v,preferably from about 50% to about 99% w/v and more preferably fromabout 80% to about 95% w/v. In a preferred embodiment, the solvent is amixture of ethanol, propylene glycol and water, more preferably fromabout 10% to about 50% w/v ethanol, from about 10% to about 70% w/vpropylene glycol and from about 10% to about 50% w/v water and even morepreferably about 28% w/v ethanol, about 47% w/v propylene glycol andabout 19% w/v water.

In one embodiment, the present invention is directed to a compositionfor the treatment of alopecia comprising a compound that binds FK506binding protein 4.

In another embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising a compound thatbinds FK506 binding protein 4 and one or more additional active agentsselected from the group consisting of minoxidil, cyclosporine A, and acombination thereof.

In another embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising a compound offormula (I) or a pharmaceutically acceptable salt or ester thereof,minoxidil and cyclosporine A,

wherein R1 is COOH, a methoxy, a phenyl, a benzyl, a substituted phenylor a substituted benzyl.

In a preferred embodiment the substituted phenyl and substituted benzylof the compound of formula (I) are each individually substituted with analkyl group, a methoxy group or a halogen.

In a more preferred embodiment the compound of formula (I) is selectedfrom the group consisting of (“RT175”), (“RT1061”), (“RT1062”) and(“RT1063”) and a pharmaceutically acceptable salt or ester thereof.

In a most preferred embodiment the compound of formula (I) is RT175.

In another embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising a compound offormula (II) or a pharmaceutically acceptable salt or ester thereof,minoxidil and cyclosporine A.

In another more preferred aspect, the compositions of the presentinvention further comprise one or more excipients selected from thegroup consisting of urea, polyoxyl 40 stearate, a carbomer, cetylalcohol, glyceryl monostearate, mineral oil, ethanol, propylene glycol,polyglycol 300, citric acid, sodium phosphate dibasic, stearyl alcohol,isopropyl myristate, sodium hydroxide, petroleum jelly, xanthan gum,white petrolatum, sorbitol solution, cetearyl alcohol, ceteareth-20,simethicone, sodium benzoate, glyceryl monostearate, polyethylene glycolmonostearate, sorbic acid, butylated hydroxytoluene and water.

In a preferred embodiment the one or more excipients are a combinationof white petrolatum, sorbitol solution, propylene glycol, cetearylalcohol, ceteareth-20, simethicone, glyceryl monostearate, polyethyleneglycol monostearate, sorbic acid and butylated hydroxytoluene.

In another preferred embodiment, the one or more excipients are acombination of urea, polyoxyl 40 stearate, propylene glycol, polyglycol300 (Medibase C available from Medisca), citric acid, sodium phosphatedibasic, cetyl alcohol, stearyl alcohol, isopropyl myristate, sodiumbenzoate and water.

In a more preferred embodiment, the one or more excipients are acombination of about 1,200 grams of urea, about 103 grams polyoxyl 40stearate, about 63 milliliters of propylene glycol, about 47 millilitersof polyglycol 300, about 1 gram of citric acid, about 2 grams of sodiumphosphate, about 94 grams of cetyl alcohol, about 200 grams of stearylalcohol, about 219 grams of isopropyl myristate, about 3 grams of sodiumbenzoate and about 1,000 to about 1,500 milliliters of water.

In a preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration from about 1% to about 10% w/v, cyclosporine A at aconcentration from about 0.01% to about 1% w/v and RT175 at aconcentration from about 0.000001% to about 0.0001% w/v.

In another preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration from about 1% to about 10% w/v, cyclosporine A at aconcentration from about 0.01% to about 1% w/v, RT175 or apharmaceutically acceptable salt or ester thereof at a concentrationfrom about 0.000001% to about 0.0001% w/v, ethanol at a concentrationfrom about 10% to about 50% w/v, propylene glycol at a concentrationfrom about 10% to about 70% w/v and water at a concentration from about10% to about 50% w/v.

In a more preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v and RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v.

In another more preferred embodiment, the present invention is directedto a composition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v, RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v, ethanol at aconcentration of about 28% w/v, propylene glycol at a concentration ofabout 47% w/v and water at a concentration of about 19% w/v.

In another embodiment, the present invention is directed to a method oftreating alopecia comprising topically administering to a human in needthereof an effective amount of a composition of the present invention.

In a preferred embodiment, the present invention is directed to a methodof treating androgenic alopecia comprising topically administering to ahuman in need thereof an effective amount of a composition of thepresent invention.

In another embodiment, the present invention is directed to a method oftreating chemotherapy-induced alopecia, including permanentchemotherapy-induced alopecia, comprising topically administering to ahuman in need thereof an effective amount of a composition of thepresent invention.

In another embodiment, the present invention is directed to a method oftreating alopecia in a human in need thereof comprising topicallyadministering concurrently or sequentially minoxidil, cyclosporine A anda compound that binds FK506 binding protein 4.

In a preferred embodiment, the human in need of alopecia treatmentsuffers from achromotrichia and the method provides regrowth ofpigmented hair, preferably the achromotrichia is due to aging.

In another embodiment, the present invention is directed to a method ofenhancing facial hair growth (including but not limited to eye brows)comprising topically administering to a human in need thereof aneffective amount of the compositions of the present invention.

In a preferred embodiment, the present invention is directed to a methodof treating alopecia in a human in need thereof comprising topicallyadministering concurrently or sequentially minoxidil at a concentrationfrom about 1% to about 10% w/v, cyclosporine A at a concentration fromabout 0.01% to about 1% w/v and RT175 or a pharmaceutically acceptablesalt or ester thereof at a concentration from about 0.000001% to about0.0001% w/v.

In a more preferred embodiment, the present invention is directed to amethod of treating alopecia in a human in need thereof comprisingtopically administering concurrently or sequentially minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v and RT175 or a pharmaceutically acceptable salt or esterthereof at a concentration of about 0.000012% w/v.

In another embodiment the present invention is directed to a method oftreating alopecia in a human in need thereof comprising the steps of:

(i) administering fractional laser treatment to an affected area of ahuman; and

(ii) topically administering a composition of the invention,

wherein the steps can be in any order.

In another embodiment the present invention is directed to a method oftreating alopecia in a human in need thereof comprising the steps of:

(i) pretreating an area affected with alopecia with 40% w/v urea; and

(ii) topically administering a composition of the invention. In anotherembodiment, the present invention is directed to a method of treatingalopecia comprising topically administering concomitantly orsequentially a compound that binds FK506 binding protein 4 and at leastone compound selected from the group consisting of minoxidil andcyclosporine A.

In another embodiment, the present invention is directed to a method oftreating alopecia comprising topically administering concomitantly orsequentially a compound that binds FK506 binding protein 4, minoxidiland cyclosporine A.

In a preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v and RT175 or a pharmaceutically acceptable salt or esterthereof, an analog thereof or a derivative thereof at a concentration ofabout 0.000012% w/v.

In another preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v, RT175 or a pharmaceutically acceptable salt or esterthereof, an analog thereof or a derivative thereof at a concentration ofabout 0.000012% w/v, ethanol at a concentration of about 28% w/v,propylene glycol at a concentration of about 47% w/v and water at aconcentration of about 19% w/v.

In another preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil at aconcentration of about 5% w/v, cyclosporine A at a concentration ofabout 0.12% w/v, RT175 or a pharmaceutically acceptable salt or esterthereof, an analog thereof or a derivative thereof at a concentration ofabout 0.000012% w/v, and one or more excipients selected from the groupconsisting of urea, a carbomer, cetyl alcohol, glyceryl monostearate,mineral oil, propylene glycol, sodium hydroxide, petroleum jelly,xanthan gum and water, in a preferred embodiment the urea is at aconcentration of about 40% w/v.

In another preferred embodiment, the present invention is directed to acomposition for the treatment of alopecia comprising minoxidil,cyclosporine A, RT175 or a pharmaceutically acceptable salt or esterthereof, an analog thereof or a derivative thereof and one or moreexcipients selected from the group consisting of urea, a carbomer, cetylalcohol, glyceryl monostearate, mineral oil, propylene glycol, sodiumhydroxide, petroleum jelly, xanthan gum and water. In this preferredembodiment the urea is at a concentration of about 40% w/v.

The formulating of the immediate above preferred embodiment may includethe following:

-   -   weigh out 100 g 40% urea cream comprising X-viate™ 40% cream: 40        g urea (w/v), carbopol, cetyl alcohol, glyceryl monostearate,        light mineral oil, propylene glycol, water, sodium hydroxide,        white petrolatum and xanthum gum; or emollient cream #1: 40 g        urea plus 60 g emollient cream #1 (polyoxyl 40 stearate (103.125        g/L), propylene glycols (62.2 ml/L), polyglycol 300MW liquid        (46.875 ml/L), citric acid-anhydrous (1.1 g/L), sodium phosphate        dibasic (2.04 g/L), cetyl alcohol wax (93.75 g/L), stearyl        alcohol wax (200 g/L), isopropyl myristate (218.75/L), dd H₂O to        1 L; sodium benzoate (3.125 g/L)); add 5 g minoxidil powder and        then add 120 mg cyclosporine A powder or 120 mg cyclosporine A        dissolved in 2 ml propylene glycol; followed by adding 50 μl of        a 1M solution RT175 (>120 ng/ml final); and then mixing the        resultant composition and loading it into 20 ml syringes or into        tubes for dispensing.

EXAMPLES Example 1. Punch Biopsies of Mouse Dorsal Skin TreatedTopically with RT175

Mice underwent 5 millimeter, full thickness, punch biopsies of theirdorsal skin. Mice were then treated topically with 120 nanograms (“ng”)of RT175, daily.

5 days after punch biopsy healing of the wound site is clearlyaccelerated in mice receiving RT175 over vehicle. See FIG. 1. 9 daysafter punch biopsy 80% of the RT175 treated wounds are completelyclosed. The other 20% of RT175 treated wounds closed within 18 hours.Additionally, 9 days after punch biopsy the RT175 treated woundsdisplayed robust hair regeneration, with intact follicles even in themiddle of the wound. See FIG. 1. By comparison, the vehicle treatedtissue has yet to regenerate follicles in the center of the wound site.

Example 2. Scalp Closure Following Craniotomy in Rats Treated Topicallywith RT175

Rats which had undergone brain surgery to test RT175 mediatedregeneration of the brain following stroke. Following surgery, the scalpwas closed. Those rats which had received RT175 treatment achieved fullscalp closure. See FIG. 2(a). However, those rats which did not receiveRT175 treatment did not achieve scalp closure. See FIG. 2(b).

Example 3. Skin Lesions in Pig Treated Topically with RT175

A single pig underwent bilateral, slit thickness, skin surgery leavinglesions on opposite sides of the pig. One of these lesions was thentreated topically with 120 ng of RT175, daily. The other lesion wastreated with vehicle only. The lesion treated with RT175, and not thelesion treated with vehicle only, exhibited early and transientappearance of granulation tissue, followed by rapid revascularization,invasion of newly formed skin from wound edges, acceleratedrepigmentation and regeneration of hair. See FIG. 3 and compare 3(b) to3(a). The de novo hair growth is consistent with the regeneration of theupper hair follicles, most of which would have been removed as themicrotome sliced through the skin in creating the split thicknesswounds.

Example 4. 59-Year-Old Male with Androgenic Alopecia Treated Topicallywith RT175 and RT175/Minoxidil

A 59-year-old white with androgenic alopecia was treated twice dailywith 1 milliliter of a composition containing 120 nanograms/milliliter(“ng/mL”) of RT175 in 30% w/v ethanol, 50% w/v propylene glycol and 20%w/v water. The composition was applied directly to the bald skin on thecrown of the head once in the morning after showering and once beforegoing to bed. Baseline hair distribution is shown in FIG. 4(a).

FIG. 4(b) shows an absence of hair growth 4 months after RT175treatment. The subject went on treatment holiday for 1 month, beforebeginning treatment with 120 ng/mL of RT175 and 5% w/v minoxidil in 30%w/v ethanol, 50% w/v propylene glycol and 20% w/v water. FIG. 4(c)depicts the subjects scalp before initiation of the second round oftreatment. FIG. 4(d) shows the failure of the combination of RT175 andminoxidil to induce new hair growth.

Example 5. 57-Year-Old Male with Androgenic Alopecia Treated Topicallywith RT175/Minoxidil/Cyclosporine A

A 57-year-old male with androgenic alopecia was treated twice daily with1 milliliter of a composition containing 120 ng/mL of RT175 (0.000012%w/v), 1.2 milligrams per milliliter (“mg/mL”) of cyclosporine A (0.12%w/v) and 5% w/v minoxidil in about 28% w/v ethanol, about 47% w/vpropylene glycol, and about 19% w/v water. The composition was applieddirectly to the bald skin on the crown of the head once in the morningafter showering and once before going to bed. The extent of hair loss isdepicted in FIG. 5 in the panel at the upper left (baseline).

FIG. 5 upper right panel shows an unexpected and extensive hair growthafter 21 days. FIG. 5 middle left panel shows continued hair growthafter 28 days. FIG. 5 middle right panel show continued hair growthafter 35 days and lower left panel shows additional hair growth at 42days. These results are surprising in light of and in stark contrast tothe findings in Example 4 that RT175 alone or in combination withminoxidil did not induce new hair growth in a male suffering fromandrogenic alopecia.

Example 6. 62-Year-Old Female with Androgenic Alopecia Treated Topicallywith RT175/Minoxidil/Cyclosporine A

A 62-year-old female with androgenic alopecia was treated twice dailywith 1 milliliter of a composition containing 120 ng/mL of RT175(0.000012% w/v), 1.2 milligrams per milliliter (“mg/mL”) of cyclosporineA (0.12% w/v) and 5% w/v minoxidil in about 28% w/v ethanol, about 47%w/v propylene glycol, and about 19% w/v water. The composition wasapplied directly to the bald skin on the crown of the head once in themorning after showering and once before going to bed. The extent of hairloss is depicted in FIG. 6a . (baseline).

FIG. 6b . shows an unexpected and extensive hair growth after 12 weeks.These results are surprising in light of and in stark contrast to thefindings in Example 4 that RT175 alone or in combination with minoxidildid not induce new hair growth in individuals suffering from androgenicalopecia.

Example 7. 57-Year-Old Male with Androgenic Alopecia and AchromotrichiaTreated Topically with RT175/Minoxidil/Cyclosporine A Displays Regrowthof Pigmented Hair

A 57-year-old male with androgenic alopecia and who had already gonethrough achromotrichia (natural loss of pigmentation in the hairfollicle and hair shaft) was treated twice daily continuously for 20weeks with 1 milliliter of a composition containing 120 ng/mL of RT175(0.000012% w/v), 1.2 milligrams per milliliter (“mg/mL”) of cyclosporineA (0.12% w/v) and 5% w/v minoxidil in about 28% w/v ethanol, about 47%w/v propylene glycol, and about 19% w/v water. The composition wasinitially applied directly to the bald skin at the crown of the headonce after showering in the morning and once before going to bed. Thecomposition continued to be applied after robust hair growth. At 5weeks, the subject began growing pigmented hair in the area of the headwhere hair loss had occurred. See FIG. 7a . The new pigmented hair ofthe subject was of a similar color as the subject's hair prior to thesubject undergoing achromotrichia. The hair outside of the treated areadid not develop pigment. By the 20^(th) week of treatment, there wasextensive growth of pigmented hair over the totality of the treatedarea. See FIG. 7b . By 1 year, there was a confluence of pigmented hairover the totality of the treated area. See FIG. 7c . This resultdemonstrates that the composition of the invention not only regrows hairin patients with androgenic alopecia but also regrows pigmented hair inpatients with androgenic alopecia who have undergone achromotrichia dueto aging.

Example 8. 57-Year-Old Male with Chemotherapy-Induced Alopecia TreatedTopically with RT175/Minoxidil/Cyclosporine A Displays Regrowth Hair

A 57-year-old male with hair loss following chemotherapy treated twicedaily with 1 milliliter of a composition containing 120 ng/mL of RT175(0.000012% w/v), 1.2 milligrams per milliliter (“mg/mL”) of cyclosporineA (0.12% w/v) and 5% w/v minoxidil in about 28% w/v ethanol, about 47%w/v propylene glycol, and about 19% w/v water. The composition wasapplied directly to the bald skin on the crown of the head once in themorning after showering and once before going to bed. The extent of hairloss is depicted in FIG. 8a . (baseline).

FIG. 8b . shows an unexpected and extensive hair growth after 21 days.These results are surprising in light of and in stark contrast to thefindings in Example 4 that RT175 alone or in combination with minoxidildid not induce new hair growth in a male suffering from androgenicalopecia.

Example 9. Prophetic Combination Treatment of Alopecia with FractionalLaser Treatment and Topical Minoxidil, Cyclosporine A or RT175 Method

Subject 1, a 50-year-old male, was subjected to topical administrationof RT175, once in the morning after showering and once before going tobed for 60 days. After 30 days-post treatment Subject 1 had notexperienced significant hair regrowth. 60 days-post RT175 treatmentSubject 1 was subjected to Fraxel® fractional laser treatment of areasof the scalp affected by alopecia using standard Fraxel® protocol. 45day-post Fraxel® treatment Subject 1 had not experienced substantialhair regrowth. 7 days later, Subject 1 was retreated with Fraxel®followed 8 hours later by topical treatment with a compositioncontaining RT175. Subject 1 then continued topical treatment with theRT175 composition for 60 days.

Results

On day 21-post Fraxel®/RT175 treatment, subject 1 began regrowing hairin areas of the scalp affected by alopecia. This regrowth continuedthrough the end of the treatment on day 90.

Example 10. 24-Year-Old Male with Inadequate Facial Hair Growth TreatedTopically with RT175/Minoxidil/Cyclosporine A Displays Enhanced Growthof Facial Hair

A 24 year old male with inadequate facial hair growth was treated twicedaily for three weeks with 0.4 milliliter of a composition containing120 ng/mL of RT175 (0.000012% w/v), 1.2 milligrams per milliliter(“mg/mL”) of cyclosporine A (0.12% w/v) and 5% w/v minoxidil inVanicream® (Vanicream is a registered trademark of Pharmaceuticalspecialties, Inc. and contains white petrolatum, sorbitol solution,propylene glycol, cetearyl alcohol, ceteareth-20, simethicone, glycerylmonostearate, polyethylene glycol monostearate, sorbic acid andbutylated hydroxytoluene). FIG. 9, upper panels, shows baseline facialhair. FIG. 9, lower panels, shows facial hair following three weeks oftwice daily treatment with RT175/cyclosporine A/minoxidil.

Example 11. 45-Year-Old Female with Inadequate Eyebrow Growth TreatedTopically with RT175/Minoxidil/Cyclosporine A Displays Enhanced Growthof Eyebrows

A 45-year-old female with inadequate eyebrow growth was treated twicedaily for twelve weeks with 0.4 milliliter of a composition containing120 ng/mL of RT175 (0.000012% w/v), 1.2 milligrams per milliliter(“mg/mL”) of cyclosporine A (0.12% w/v) and 5% w/v minoxidil inVanicream®. FIG. 10a . shows baseline eyebrows. FIG. 10b-d . showseyebrows following 3, 9 and 12 weeks of twice daily treatment withRT175/cyclosporine A/minoxidil.

Example 12. Treatment of Dermabrasion Lesions of Dorsal Skin of Micewith RT175 Analog, RT1061

Retired ICR breeder mice of both sexes with weight between 45 and 50grams were shaved and rasped to induce dermabrasion lesions. The animalswere then randomly assigned (5 per treatment arm) to one of two groupsto receive either 5 μL of 100 nanomolar RT1061 or a vehicle control.RT1061 or vehicle control was applied topically to the lesion site onceeach day including immediately after dermabrasion.

Results of the study can be seen in FIG. 11. As shown, treatment withRT1061 (panels b., d. and f.) resulted in rapid hair growth and rapidregeneration of rasped skin. In contrast, treatment with the controlvehicle resulted in scab formation and no hair regrowth 10 days afterrasping. See FIG. 11, panels a., c. and e.

What is claimed is:
 1. A method of treating chemotherapy-inducedalopecia comprising topically administering to a human in need thereofan effective amount of a composition comprising a compound that bindsFK506 binding protein
 4. 2. The method of claim 1 wherein thecomposition further comprises one or more additional active agentsselected from the group consisting of minoxidil, cyclosporine A, and acombination thereof.
 3. The method of claim 1, wherein thechemotherapy-induced alopecia is permanent.
 4. A method of treatingchemotherapy-induced alopecia comprising topically administering to ahuman in need thereof an effective amount of a composition comprising: acompound that binds FK506 binding protein 4; minoxidil; and cyclosporineA.
 5. The method of claim 4, wherein the chemotherapy-induced alopeciais permanent.
 6. A method of treating chemotherapy-induced alopeciacomprising topically administering to a human in need thereof aneffective amount of a composition for the treatment of alopeciacomprising a compound of formula (I)

or a pharmaceutically acceptable salt or ester thereof, minoxidil andcyclosporine A, wherein R¹ is selected from the group consisting ofCOOH, a methoxy, a phenyl, a benzyl, a substituted phenyl and asubstituted benzyl.
 7. The method of claim 6, wherein the substitutedphenyl and the substituted benzyl are each individually substituted withan alkyl group, a methoxy group or a halogen.
 8. The method of claim 6,wherein the compound of formula (I) is selected from the groupconsisting of

and a pharmaceutically acceptable salt or ester thereof.
 9. The methodof claim 6, wherein the chemotherapy-induced alopecia is permanent. 10.A method of treating chemotherapy-induced alopecia comprising topicallyadministering to a human in need thereof an effective amount of acomposition comprising a compound of formula (II)

or a pharmaceutically acceptable salt or ester thereof, minoxidil andcyclosporine A.
 11. The method of claim 10, wherein thechemotherapy-induced alopecia is permanent.
 12. The method of claim 10,wherein the composition further comprises one or more excipientsselected from the group consisting of urea, polyoxyl 40 stearate, acarbomer, cetyl alcohol, glyceryl monostearate, mineral oil, ethanol,propylene glycol, polyglycol 300, citric acid, sodium phosphate dibasic,stearyl alcohol, isopropyl myristate, sodium hydroxide, petroleum jelly,xanthan gum, white petrolatum, sorbitol solution, cetearyl alcohol,ceteareth-20, simethicone, sodium benzoate, glyceryl monostearate,polyethylene glycol monostearate, sorbic acid, butylated hydroxytolueneand water.
 13. The method of claim 10, wherein minoxidil is at aconcentration from about 1% to about 10% w/v, cyclosporine A is at aconcentration from about 0.01% to about 1% w/v and the compound offormula (II) or a pharmaceutically acceptable salt or ester thereof isat a concentration from about 0.000001% to about 0.0001% w/v, whereinw/v denotes weight by volume.
 14. The method of claim 10, furthercomprising ethanol at a concentration from about 10% to about 50% w/v,propylene glycol at a concentration from about 10% to about 70% w/v andwater at a concentration from about 10% to about 50% w/v.
 15. The methodof claim 10, further comprising 40% w/v urea.
 16. The method of claim10, further comprising polyoxyl 40 stearate, citric acid, polyglycol300, sodium phosphate, cetyl alcohol, stearyl alcohol, isopropylmyristate and sodium benzoate.
 17. The method of claim 10, whereinminoxidil is at a concentration of about 5% w/v, cyclosporine A is at aconcentration of about 0.12% w/v and the compound of formula (II) or apharmaceutically acceptable salt or ester thereof is at a concentrationof about 0.000012% w/v.
 18. The method of claim 10, further comprisingethanol at a concentration of about 28% w/v, propylene glycol at aconcentration of about 47% w/v and water at a concentration of about 19%w/v.
 19. A method of treating alopecia comprising topicallyadministering to a female human in need thereof an effective amount of acomposition comprising a compound that binds FK506 binding protein 4,minoxidil; and cyclosporine A.
 20. A method of enhancing eyebrow growthcomprising topically administering to a human in need thereof aneffective amount of a composition comprising a compound that binds FK506binding protein 4, minoxidil; and cyclosporine A.